surgical-placement-catheters-for-long-term-cardiovascular-exercise

Surgical Placement of Catheters for Long-term Cardiovascular Exercise Testing in Swine

This protocol describes the surgical procedure to chronically instrument swine and the procedure to exercise swine on a motor-driven treadmill. Early ...

preparation-of-human-myocardial-tissue-for-long-term-cultivation

Preparation of Human Myocardial Tissue for Long-Term Cultivation

Cardiomyocyte cultivation has seen a vast number of developments, ranging from two-dimensional (2D) cell cultivation to iPSC derived organoids. In 2019, ...

Erasmus MC

University Hospital, LMU Munich

German Center for Cardiovascular Research (DZHK), Partner Site Munich, Munich Heart Alliance (MHA)

University Medical Center Rotterdam

Pulmonary hypertension (PH) as a result of pulmonary vein stenosis (PVS) is extremely difficult to treat. The ideal therapy should not target the high-pressure/low-flow (HP/LF) vasculature that drains into stenotic veins, but only the high-pressure/high-flow (HP/HF) vasculature draining into unaffected pulmonary veins, reducing vascular resistance and pressure without risk of pulmonary oedema. We aimed to assess the activity of the nitric oxide (NO) pathway in PVS during the development of PH, and investigate whether interventions in the NO pathway differentially affect vasodilation in the HP/HF vs. HP/LF territories. Swine underwent pulmonary vein banding (PVB;  = 7) or sham surgery ( = 6) and were chronically instrumented to assess progression of PH. Pulmonary sensitivity to exogenous NO (sodium nitroprusside, SNP) and the contribution of endogenous NO were assessed bi-weekly. The pulmonary vasodilator response to phosphodiesterase-5 (PDE5) inhibition was assessed 12 weeks after PVB or sham surgery. After sacrifice, 12 weeks post-surgery, interventions in the NO pathway on pulmonary small arteries isolated from HP/LF and HP/HF territories were further investigated. There were no differences in the in vivo pulmonary vasodilator response to SNP and the pulmonary vasoconstrictor response to endothelial nitric oxide synthase (eNOS) inhibition up to 8 weeks after PVB as compared to the sham group. However, at 10 and 12 weeks post-PVB, the in vivo pulmonary vasodilation in response to SNP was larger in the PVB group. Similarly, the vasoconstriction to eNOS inhibition was larger in the PVB group, particularly during exercise, while pulmonary vasodilation in response to PDE5 inhibition was larger in the PVB group both at rest and during exercise. In isolated pulmonary small arteries, sensitivity to NO donor SNP was similar in PVB vs. sham groups irrespective of HP/LF and HP/HF, while sensitivity to the PDE5 inhibitor sildenafil was lower in PVB HP/HF and sensitivity to bradykinin was lower in PVB HP/LF. In conclusion, both NO availability and sensitivity were increased in the PVB group. The increased nitric oxide sensitivity was not the result of a decreased PDE5 activity, as PDE5 activity was even increased. Some vasodilators differentially effect HP/HF vs. HP/LF vasculature.

Intervening with the Nitric Oxide Pathway to Alleviate Pulmonary Hypertension in Pulmonary Vein Stenosis.

Activation of both adenosine A and A receptors (AR) contributes to coronary vasodilation. We previously demonstrated that uridine adenosine tetraphosphate (UpA) is a novel vasodilator in the porcine coronary microcirculation, acting mainly on AR in smooth muscle cells (SMC). We further investigated whether activation of AR is involved in UpA-mediated coronary SMC relaxation. Both AR and AR may stimulate HO production leading to activation of K channels in SMCs, we also studied the involvement of HO and K channels in UpA-mediated effect. Coronary small arteries dissected from the apex of porcine hearts were mounted on wire myograph for UpA concentration responses. UpA-induced coronary SMC relaxation was attenuated by AR but not AR antagonism or non-selective P2R antagonism, despite greater endogenous AR expression vs. AR in both coronary small arteries and primary cultured coronary SMCs. Moreover, UpA-induced coronary SMC relaxation was blunted by HO catabolism. This effect was not altered by K channel blockade. Combination of HO catabolism and AR antagonism attenuated UpA-induced coronary SMC relaxation to the similar extent as AR antagonism alone. Collectively, UpA-induced porcine coronary SMC relaxation is mediated by activation of AR-HO pathway. This process does not involve AR, P2R or K channels.

Activation of adenosine A but not A receptors is involved in uridine adenosine tetraphosphate-induced porcine coronary smooth muscle relaxation.

Coronary microvascular dysfunction (CMD) is commonly present in patients with metabolic derangements and is increasingly recognized as an important contributor to myocardial ischaemia, both in the presence and absence of epicardial coronary atherosclerosis. The latter condition is termed 'ischaemia and no obstructive coronary artery disease' (INOCA). Notwithstanding the high prevalence of INOCA, effective treatment remains elusive. Although to date there is no animal model for INOCA, animal models of CMD, one of the hallmarks of INOCA, offer excellent test models for enhancing our understanding of the pathophysiology of CMD and for investigating novel therapies. This article presents an overview of currently available experimental models of CMD-with an emphasis on metabolic derangements as risk factors-in dogs, swine, rabbits, rats, and mice. In all available animal models, metabolic derangements are most often induced by a high-fat diet (HFD) and/or diabetes mellitus via injection of alloxan or streptozotocin, but there is also a wide variety of spontaneous as well as transgenic animal models which develop metabolic derangements. Depending on the number, severity, and duration of exposure to risk factors-all these animal models show perturbations in coronary microvascular (endothelial) function and structure, similar to what has been observed in patients with INOCA and comorbid conditions. The use of these animal models will be instrumental in identifying novel therapeutic targets and for the subsequent development and testing of novel therapeutic interventions to combat ischaemic heart disease, the number one cause of death worldwide.

Experimental animal models of coronary microvascular dysfunction.

The worldwide prevalence of diabetes mellitus and obesity is rapidly increasing not only in adults but also in children and adolescents. Diabetes is associated with macrovascular complications increasing the risk for cardiovascular disease and stroke, as well as microvascular complications leading to diabetic nephropathy, retinopathy and neuropathy. Animal models are essential for studying disease mechanisms and for developing and testing diagnostic procedures and therapeutic strategies. Rodent models are most widely used but have limitations in translational research. Porcine models have the potential to bridge the gap between basic studies and clinical trials in human patients. This article provides an overview of concepts for the development of porcine models for diabetes and obesity research, with a focus on genetically engineered models. Diabetes-associated ocular, cardiovascular and renal alterations observed in diabetic pig models are summarized and their similarities with complications in diabetic patients are discussed. Systematic multi-organ biobanking of porcine models of diabetes and obesity and molecular profiling of representative tissue samples on different levels, e.g., on the transcriptome, proteome, or metabolome level, is proposed as a strategy for discovering tissue-specific pathomechanisms and their molecular key drivers using systems biology tools. This is exemplified by a recent study providing multi-omics insights into functional changes of the liver in a transgenic pig model for insulin-deficient diabetes mellitus. Collectively, these approaches will provide a better understanding of organ crosstalk in diabetes mellitus and eventually reveal new molecular targets for the prevention, early diagnosis and treatment of diabetes mellitus and its associated complications.

Porcine models for studying complications and organ crosstalk in diabetes mellitus.

 Exogenous melatonin has been reported to be beneficial in the treatment of pulmonary hypertension (PH) in animal models. Multiple mechanisms are involved, with melatonin exerting anti-oxidant and anti-inflammatory effects, as well as inducing vasodilation and cardio-protection. However, endogenous levels of melatonin in treatment-naïve patients with PH and their clinical significance are still unknown. Plasma levels of endogenous melatonin were measured by liquid chromatography-tandem mass spectrometry in PH patients ( = 64, 43 pulmonary arterial hypertension (PAH) and 21 chronic thromboembolic PH (CTEPH)) and healthy controls ( = 111). Melatonin levels were higher in PH, PAH, and CTEPH patients when compared with controls (Median 118.7 (IQR 108.2-139.9), 118.9 (109.3-147.7), 118.3 (106.8-130.1) versus 108.0 (102.3-115.2) pM, respectively,  all < 0.001). The mortality was 26% (11/43) in the PAH subgroup during a long-term follow-up of 42 (IQR: 32-58) months. Kaplan-Meier analysis showed that, in the PAH subgroup, patients with melatonin levels in the 1st quartile (<109.3 pM) had a worse survival than those in quartile 2-4 (Mean survival times were 46 (95% CI: 30-65) versus 68 (58-77) months, Log-rank,  = 0.026) with an increased hazard ratio of 3.5 (95% CI: 1.1-11.6,  = 0.038). Endogenous melatonin was increased in treatment-naïve patients with PH, and lower levels of melatonin were associated with worse long-term survival in patient with PAH.

Lower Plasma Melatonin Levels Predict Worse Long-Term Survival in Pulmonary Arterial Hypertension.

The prevalence of diabetic metabolic derangement (DMetD) has increased dramatically over the last decades. Although there is increasing evidence that DMetD is associated with cardiac dysfunction, the early DMetD-induced myocardial alterations remain incompletely understood. Here, we studied early DMetD-related cardiac changes in a clinically relevant large animal model. DMetD was established in adult male Göttingen miniswine by streptozotocin injections and a high-fat, high-sugar diet, while control animals remained on normal pig chow. Five months later left ventricular (LV) function was assessed by echocardiography and hemodynamic measurements, followed by comprehensive biochemical, molecular and histological analyses. Robust DMetD developed, evidenced by hyperglycemia, hypercholesterolemia and hypertriglyceridemia. DMetD resulted in altered LV nitroso-redox balance, increased superoxide production-principally due to endothelial nitric oxide synthase (eNOS) uncoupling-reduced nitric oxide (NO) production, alterations in myocardial gene-expression-particularly genes related to glucose and fatty acid metabolism-and mitochondrial dysfunction. These abnormalities were accompanied by increased passive force of isolated cardiomyocytes, and impaired LV diastolic function, evidenced by reduced LV peak untwist velocity and increased E/e'. However, LV weight, volume, collagen content, and cardiomyocyte cross-sectional area were unchanged at this stage of DMetD. In conclusion, DMetD, in a clinically relevant large-animal model results in myocardial oxidative stress, eNOS uncoupling and reduced NO production, together with an altered metabolic gene expression profile and mitochondrial dysfunction. These molecular alterations are associated with stiffening of the cardiomyocytes and early diastolic dysfunction before any structural cardiac remodeling occurs. Therapies should be directed to ameliorate these early DMetD-induced myocardial changes to prevent the development of overt cardiac failure.

Cellular, mitochondrial and molecular alterations associate with early left ventricular diastolic dysfunction in a porcine model of diabetic metabolic derangement.

It is well known that high von Willebrand factor (VWF) and factor VIII (FVIII) levels are associated with an increased risk of cardiovascular disease. It is still debated whether VWF and FVIII are biomarkers of endothelial dysfunction and atherosclerosis or whether they have a direct causative role. Therefore, we aimed to unravel the pathophysiological pathways of increased VWF and FVIII levels associated with cardiovascular risk factors. First, we performed a randomized controlled trial in 34 Göttingen miniswine. Diabetes mellitus (DM) was induced with streptozotocin and hypercholesterolemia (HC) via a high-fat diet in 18 swine (DM + HC), while 16 healthy swine served as controls. After 5 months of follow-up, FVIII activity (FVIII:C) was significantly higher in DM + HC swine (5.85 IU/mL [5.00-6.81]) compared with controls (4.57 [3.76-5.40],  = 0.010), whereas VWF antigen (VWF:Ag) was similar (respectively 0.34 IU/mL [0.28-0.39] vs. 0.34 [0.31-0.38],  = 0.644). DM + HC swine had no endothelial dysfunction or atherosclerosis during this short-term follow-up. Subsequently, we performed a long-term (15 months) longitudinal cohort study in 10 Landrace-Yorkshire swine, in five of which HC and in five combined DM + HC were induced. VWF:Ag was higher at 15 months compared with 9 months in HC (0.37 [0.32-0.42] vs. 0.27 [0.23-0.40],  = 0.042) and DM + HC (0.33 [0.32-0.37] vs. 0.25 [0.24-0.33],  = 0.042). Both long-term groups had endothelial dysfunction compared with controls and atherosclerosis after 15 months. In conclusion, short-term hyperglycemia and dyslipidemia increase FVIII, independent of VWF. Long-term DM and HC increase VWF via endothelial dysfunction and atherosclerosis. Therefore, VWF seems to be a biomarker for advanced cardiovascular disease.

Endothelial Dysfunction, Atherosclerosis, and Increase of von Willebrand Factor and Factor VIII: A Randomized Controlled Trial in Swine.

Chronic thrombo-embolic pulmonary hypertension (CTEPH) develops in a subset of patients after acute pulmonary embolism. In CTEPH, pulmonary vascular resistance, which is initially elevated due to the obstructions in the larger pulmonary arteries, is further increased by pulmonary microvascular remodeling. The increased afterload of the right ventricle (RV) leads to RV dilation and hypertrophy. This RV remodeling predisposes to arrhythmogenesis and RV failure. Yet, mechanisms involved in pulmonary microvascular remodeling, processes underlying the RV structural and functional adaptability in CTEPH as well as determinants of the susceptibility to arrhythmias such as atrial fibrillation in the context of CTEPH remain incompletely understood. Several large animal models with critical clinical features of human CTEPH and subsequent RV remodeling have relatively recently been developed in swine, sheep, and dogs. In this review we will discuss the current knowledge on the processes underlying development and progression of CTEPH, and on how animal models can help enlarge understanding of these processes.

Daphne Merkus

Experimental Cardiology

Erasmus MC

Surgical Placement of Catheters for Long-term Cardiovascular Exercise Testing in Swine

Preparation of Human Myocardial Tissue for Long-Term Cultivation

Daphne Merkus

.css-o250i3{font-size:18px;font-family:Open Sans,sans-serif;line-height:21.6px;}Experimental Cardiology

Erasmus MC

Surgical Placement of Catheters for Long-term Cardiovascular Exercise Testing in Swine

Preparation of Human Myocardial Tissue for Long-Term Cultivation

Experimental Cardiology